Pre-eclampsia (PE) remains one of the most common causes of adverse pregnancy outcome in developed and developing countries. The incidence of PE is substantial, about 3% to 8%1,2. PE places the obstetric patient and her baby at substantial risk of pre-term birth and perinatal mortality, and severe maternal hypertension and multi-systemic organ dysfunction and damage, including eclampsia and abruption placentae3,4. Predictive tests for pre-eclampsia early in the course of pregnancy would provide sufficient time to intervene and mitigate the risks of PE. There has been an intense interest in biomarkers for the identification of patients at risk for preeclampsia. Although clinical risk factors for pre-eclampsia are well known, these factors either singly or in combination have limited predictive values and this has led to intense search for predictive biomarkers for PE, particularly in plasma5. However, plasma-derived predictive biomarkers like the generic disease biomarkers are generally low abundance proteins and their discovery is confounded by the dominance of several high abundance proteins such as albumin and immunoglobulins. Despite much effort to eliminate or reduce these abundant proteins, circumventing these high abundance plasma proteins remains a challenge. However, the recent extraction of membrane vesicles from bodily fluids such as plasma or urine6 for biomarker discovery inadvertently resolved this challenge as removal of the high abundance plasma proteins is inherent in the extraction of membrane vesicles.
The cell sources of these circulating vesicles are likely to be diverse as many cell types are known to secrete membrane vesicles. Since these vesicles are essentially fragments of the secreting cells, they and their cargo are microcosms of their cell sources and would reflect the physiological or diseased state of the cells, making them potential sources of biomarkers for disease diagnosis or prognosis. Indeed, pregnancy-associated exosomes were reported as early as 20067. Circulating plasma vesicles are highly heterogeneous and several distinct classes of membrane vesicles have been described. They include microvesicles, ectosomes, membrane particles, exosome-like vesicles, apoptotic bodies, prostasomes, oncosomes, or exosomes, and are differentiated based on their biogenesis pathway, size, flotation density on a sucrose gradient, lipid composition, sedimentation force, and cargo content6,8,9. Presently, these vesicles are isolated by differential and/or density gradient centrifugation which rely primarily on the size or density of the vesicles. Since size and density distribution not discretely unique to each class of membrane vesicles, the present isolation techniques cannot differentiate between the different classes. Although immuno-isolation techniques using antibodies against specific membrane proteins could enhance the specificity of membrane vesicle isolation, no membrane protein has been reported to be unique to a class of membrane vesicles or to a particular cell type. For example, while tetraspanins such as CD9, CD81 have often been used as exosome-associated markers, their ubiquitous distribution over the surface membrane of many cell types suggests a generic association with membrane vesicles. Also, such immuno-isolation techniques cannot distinguish between membrane vesicles, protein complexes or soluble receptors. The lack of specific isolation technique for each class of these membrane vesicles is further exacerbated by a lack of nomenclature standard to unambiguously define each class of membrane vesicle10. It is also not clear if the present classification of vesicles describe unique entities.